Sialic acid (Sia)-containing structures in eukaryotic systems play important roles in a variety of physiological and pathological processes, including cell-cell interactions, inflammation, fertilization, viral infection, differentiation, malignancies, and cell signaling (see, e.g., Chen and Varki, ACS Chem. Biol. 2010, 5, 163-176; Traving and Schauer, Cell Mol. Life Sci. 1998, 54, 1330-1349; Schauer, Curr. Opin. Struct. Biol. 2009, 19, 507-514; Varki, Nature 2007, 446, 1023-1029). Among more than 50 different sialic acid structures that have been identified in nature, N-acetylneuraminic acid (Neu5Ac) is the most common and the most abundant sialic acid form. Sialyltransferases (EC 2.4.99.X) are key enzymes involved in the biosynthesis of these sialic acid-containing oligosaccharides and glycoconjugates (see, Harduin-Lepers, et al. Glycobiology 1995, 5, 741-758). They catalyze the transfer of a sialic acid residue from its activated sugar nucleotide donor cytidine 5′-monophosphate sialic acid (CMP-sialic acid) to an acceptor, usually a structure with a galactose (Gal), an N-acetylgalactosamine (GalNAc), an N-acetylglucosamine (GlcNAc), or another sialic acid residue. Various linkages including Siaα2-3Gal, Siaα2-6Gal/GalNAc/GlcNAc, Siaα2-8Sia, and Siaα2-9Sia can be formed. Bifunctional glycosyltransferases (SiaD) that are responsible for the formation of Neu5Ac-containing Neisseria meningitidis serogroups W-135 and Y capsular polysaccharides (CPSs) [-6Gal/Glcα1-4Neu5Acα2-]n have been grouped together with other glycosyltransferases in glycosyltransferase 4 (GT4) family in the Carbohydrate Activated enZyme (CAZy) database based on protein sequence homology (see, Bhattacharjee, et al. Can. J. Biochem. 1976, 54, 1-8; Campbell, et al. Biochem. J. 1997, 326, 929-939; Coutinho, et al. J. Mol. Biol. 2003, 328, 307-317). All other sialyltransferases reported to date have been grouped into five CAZy glycosyltransferase (GT) families (GT29, GT38, GT42, GT52, and GT80). All known eukaryotic sialyltransferases belong to a single CAZy GT29 family, while bacterial sialyltransferases are more spread out among CAZy GT families GT38, GT42, GT52, and GT80 (see, Li and Chen, Appl. Microbiol. Biotechnol. 2012, 94, 887-905; Audry, et al. Glycobiology 2011,21, 716-726).
Since bacterial sialyltransferases can be produced more easily as active forms in larger amounts in Escherichia coli expression systems and many of them have broader substrate specificities than their mammalian counterparts, they have been used as efficient catalysts in preparative and large scale synthesis of biologically important sialosides (see, Yamamoto, Mar. Drugs 2010, 8, 2781-2794; Yu and Chen, et al. Angew. Chem. Int. Ed. 2006, 45, 3938-3944). For example, multifunctional Pasteurella multocida α2-3-sialyltransferase 1 (PmST1) has been used as a powerful catalyst in the chemoenzymatic synthesis of diverse α2-3-linked sialosides (see, Yu and Chen, et al. J. Am. Chem. Soc. 2005, 127, 17618-17619). Photobacterium damselae α2-6-sialyltransferase (Pd2,6ST) has been applied in the synthesis of α2-6-linked sialosides and glycopeptides (see, Yu, Angew. Chem. Int. Ed. 2006, supra; Yamamoto, et al. Biosci. Biotechnol. Biochem. 1998, 62, 210-214; Kajihara, et al. Carbohydr. Res. 1999, 315, 137-141; Teo, et al. Adv. Synth. Catal. 2005, 347, 967-972; Yu and Chen, et al. Nat. Protoc. 2006, 1, 2485-2492). Campylobacter jejuni OH4384 α2-3/8-sialyltransferase (CstII) has been used for the synthesis of GD3 and GT1a ganglioside oligosaccharides (see, Gilbert, et al. Biol. Chem. 2000, 275, 3896-3906; Blixt, et al. Carbohydr. Res. 2005, 340, 1963-1972; Antoine, et al. Angew. Chem. Int. Ed. 2005, 44, 1350-1352; Cheng and Chen, et al. Glycobiology 2008, 18, 686-697; Yu and Chen, et al. J. Am. Chem. Soc. 2009, 131, 18467-18477).
Among sialic acid-containing biologically important sialosides, sialyl Tn antigens (Siaα2-6GalNAcα1-O-Ser/Thr) have been reported to correlate with the invasive and metastatic growth of carcinoma cells and are considered as a tumor-associated antigens for cancer vaccination development (Wu and Guo. Bioconj. Chem. 2006, 17, 1537-1544). In addition to conventional chemical methods for synthesis of sialyl Tn (STn) antigens, sialyltransferase-catalyzed glycosylation has been shown as a highly efficient approach. The present inventors previously identified recombinant Photobacterium sp. JH-ISH-224 α2-6-sialyltransferase Psp26ST(15-501)-His6 as a more suitable α2-6-sialyltransferase than Pd2,6ST for catalyzing the formation of STn antigens from N-acetylgalactosamine (GalNAc)-containing glycosides such as GalNAcα2AA, GalNAcαOSer, and GalNAcαOThr as acceptor substrates (Ding and Chen, et al. Chem. Commun. 2011, 47, 8691-8693). Nevertheless, the efficiency of Psp26ST(15-501)-His6 in sialylating α-GalNAc-terminated glycosides (Tn-antigens) is still much lower than sialylating β-galactosides. In addition, the expression level of soluble Psp2,6ST(15-501)-His6 (25 mg L−1) is not as high as Pd2,6ST (36 mg L−1; see, Sun and Chen, et al. Biotechnology Letters 2008, 30, 671-676). Sialyltransferases exhibiting high expression yield and high catalytic efficiency are needed in order to expand synthetic methodology for preparation of STn antigens and other biologically important sialosides. The present invention addresses this and other needs.